Thermococcus profundus 2-Ketoisovalerate Ferredoxin Oxidoreductase, a Key Enzyme in the Archaeal Energy-Producing Amino Acid Metabolic Pathway

• Published in: Journal of biochemistry (Tokyo) 2005, Vol.137 (1), p.101-107
• Main Authors: Ozawa, Yukiko, Nakamura, Takahiro, Kamata, Natsu, Yasujima, Daisuke, Urushiyama, Akio, Yamakura, Fumiyuki, Ohmori, Daijiro, Imai, Takeo
• Format: Article
• Language: English
• Subjects: 2-ketoglutarate ferredoxin oxidoreductase; 2-ketoisovalerate; 2-ketoisovalerate ferredoxin oxidoreductase; amino acid; Amino Acids - metabolism; Energy Metabolism - physiology; ferredoxin; hyperthermophile; indolepyruvate ferredoxin oxidoreductase; IOR; Iron-Sulfur Proteins - chemistry; Iron-Sulfur Proteins - isolation & purification; Iron-Sulfur Proteins - metabolism; Ketone Oxidoreductases - chemistry; Ketone Oxidoreductases - isolation & purification; Ketone Oxidoreductases - metabolism; KGOR; oxygen-sensitive; PFOR; Protein Subunits - chemistry; Protein Subunits - isolation & purification; Protein Subunits - metabolism; pyruvate ferredoxin oxidoreductase; resonance Raman; Thermococcus - enzymology; thiamine pyrophosphate; TPP; VOR
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/jb/mvi012

2-Ketoisovalerate ferredoxin oxidoreductase (VOR) is a key enzyme in hyperthermophiles catalyzing the coenzyme A-dependent oxidative decarboxylation of mainly aliphatic amino acid-derived 2-keto acids. The very oxygen-labile enzyme purified under anaerobic conditions from a hyperthermophilic archaeon, Thermococcus profundus, is a hetero-octamer ([alpha]{szligbeta}[gamma][delta])₂ consisting of four different subunits, [alpha] = 45,000, {szligbeta} = 31,000, [gamma] = 22,000 and [delta] = 13,000, respectively. Electron paramagnetic resonance and resonance Raman spectra of the purified enzyme indicate the presence of approximately three [4Fe-4S] clusters per [alpha]{szligbeta}[gamma][delta]-protomer, although one of the clusters has a tendency to be converted to a [3Fe-4S] form during purification. The optimal temperature for the enzyme activity is 93 ± 2°C and the cognate [4Fe-4S] ferredoxin serves as an electron acceptor of the enzyme. The purified enzyme is highly oxygen-labile (t[subscript 1/2], approximately 5 min at 25°C), and is partly protected in the presence of magnesium ions, thiamine pyrophosphate and sodium chloride (t[subscript 1/2], approximately 25 min at 25°C).